Most biologic systems show circadian rhythms probably regulated by signals from the suprachiasmatic nucleus (SCN) in the hypothalamus. Hematopoietic rhythms have been identified for different progenitor cells, marrow DNA synthesis, and various hematopoietically active cytokines. We have now identified prominent biologic rhythms for murine marrow stem cell engraftment into 100 cGy treated mice. We have also established a unique, Lin- Ho(low) Rho(low) double efflux murine stem cell separation, and identified the "R1" subpopulation of these cells as appropriate for these circadian studies. In the present grant, we plan to evaluate the role of extraneous factors, the rhythms of short-term (3 weeks) and longer-term (6, 9 or 12 months), and whether there are seasonal rhythms for engraftment. We will also evaluate whether the engraftment rhythm is based on a homing rhythm. In order to facilitate these studies and to begin to investigate molecular and biochemical mechanisms of these rhythms, we will study the temporal rhythms of engraftability and luciferase expression in Per1-luc transgenic rats (and mice). As a prerequisite for these studies, we will establish a rat stem cell purification model and a rat marrow transplantation-tracking model. This will enable us to monitor rhythms after stem cell separation and purification procedures. We will also study whether murine stem cell engraftment rhythms persist after a stem cell purification procedure. We will evaluate whether ablating the SCN abolishes these rhythms. Lastly, with availability of Per1-luc purified rat stem cells, we will study gene and protein expression using immunophenotyping and array technology. These experiments will employ congenic Ly-5.1 to Ly-5.2 or PVG RT7b to PVG RT7a (rats) transplants in 100 cGy exposed mice or rats, conditioning of rodents in light-dark boxes, in vitro progenitor clonal culture and tritiated thymidine suicide. Donor cells will be tracked using monoclonal antibodies, fluorescence in situ hybridization (FISH), and FACS analysis. Transgenic Pert-luc rats and mice will be available at University of Virginia, and the array/informatic studies done at Princeton.